Role of Na+/H+ exchanger during O2 deprivation in mouse CA1 neurons

1 Section of Respiratory Medicine, Department of Pediatrics, and 2 Department of Cellular and Molecular Physiology, Yale University School of Medicine, New Haven, Connecticut 06520 To determine the role of membrane transporters in intracellular pH (pH i ) regulation under conditions of low microenvironmental O 2 , we monitored pH i in isolated single CA1 neurons using the fluorescent indicator carboxyseminaphthorhodafluor-1 and confocal microscopy. After total O 2 deprivation or anoxia (P O 2 0 Torr), a large increase in pH i was seen in CA1 neurons in HEPES buffer, but a drop in pH i , albeit small, was observed in the presence of HCO . Ionic substitution and pharmacological experiments showed that the large anoxia-induced pH i increase in HEPES buffer was totally Na + dependent and was blocked by HOE-694, strongly suggesting the activation of the Na + /H + exchanger (NHE). Also, this pH i increase in HEPES buffer was significantly smaller in Na + /H + exchanger isoform 1 (NHE1) null mutant CA1 neurons than in wild-type neurons, demonstrating that NHE1 is responsible for part of the pH i increase following anoxia. Both chelerythrine and H-89 partly blocked, and H-7 totally eliminated, this anoxia-induced pH i increase in the absence of HCO . We conclude that 1 ) O 2 deprivation activates Na + /H + exchange by enhancing protein kinase activity and 2 ) membrane proteins, such as NHE, actively participate in regulating pH i during low-O 2 states in neurons. hippocampus; transporter; anoxia; pH; sodium-hydrogen exchanger